Continuation of the isolation and determination of amino acid sequences of those proteins present in mammalian intracellular membranes is the broad objective of this proposal. The Principal Investigator has already isolated and elucidated the amino acid sequence of the enzymatically active, water-soluble moiety of cytochrome b5 from six mammalian species. Isolation and partial determination of the primary structure of cytochrome b5 obtained from an individual human liver was also accomplished. This large molecular- weight protein was found to have both a catalytic moiety and a segment which presumably is responsible for the strong association between the cytochrome and the endoplasmic membrane. Among the specific objectives of the proposed study are the following: (1) a detailed study of the amino acid sequences of cytochrome b5 in normal and pathological human liver; (2) an attempt to characterize the primary structure of the cytochrome P-450-like heme protein unexpectedly isolated during isolation of human cytochrome b5; (3) the isolation and determination of the primary structure of the entire liver microsomal b5 molecule from the key mammalian species; (4) the examination of the cytochrome b5 sequences occurring in bovine kidney and heart membranes; (5) the continuation of collaborative elucidation studies of the primary structure of calf cytochrome b5 reductase. Information from these studies will contribute to understanding of the structure, synthesis and evolution of membranous proteins. Characterization of cytochrome b5 structure from individual human livers will provide molecular evidence for differences between individual members of the human population. Studies comparing cytochrome b5 sequences from normal cells with those isolated from neoplastic hepatocytes will provide an explanation to the presently unanswered question of whether or not protein biosynthesis is altered in neoplasia. If we are successful in characterizing the cytochrome P-450-like heme protein, we will then know its molecular properties which should provide information about its function and structure, as well as its relationship to the drug-induced cytochrome P-450.